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- EMDB-11526: Cryo-EM structure of the SARS-CoV-2 spike protein bound to neutra... -

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Basic information

Entry
Database: EMDB / ID: EMD-11526
TitleCryo-EM structure of the SARS-CoV-2 spike protein bound to neutralizing nanobodies (Ty1)
Map dataComposite map of prefusion SARS-CoV-2 spike glycoprotein with a single receptor binding domain up. The spike protein is bound with three neutralizing nanobodies.
Sample
  • Complex: Ternary complex between Nanobody Ty1 and Spike.
    • Complex: Nanobody strongly neutralising and blocks ACE2 binding.
      • Protein or peptide: Spike glycoproteinSpike protein
    • Complex: Nanobody Ty1
      • Protein or peptide: Nanobody Ty1
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Function / homology
Function and homology information


Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / host cell endoplasmic reticulum-Golgi intermediate compartment membrane ...Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / entry receptor-mediated virion attachment to host cell / receptor-mediated endocytosis of virus by host cell / Attachment and Entry / membrane fusion / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / receptor ligand activity / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / virion attachment to host cell / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / virion membrane / identical protein binding / plasma membrane
Similarity search - Function
Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like ...Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Betacoronavirus-like spike glycoprotein S1, N-terminal / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 2 (HR2) region profile. / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 1 (HR1) region profile. / Spike glycoprotein S2 superfamily, coronavirus / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2 / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal
Similarity search - Domain/homology
Biological speciesSevere acute respiratory syndrome coronavirus 2 / Vicugna pacos (alpaca)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.93 Å
AuthorsHallberg BM / Das H
Citation
Journal: Nat Commun / Year: 2020
Title: An alpaca nanobody neutralizes SARS-CoV-2 by blocking receptor interaction.
Authors: Leo Hanke / Laura Vidakovics Perez / Daniel J Sheward / Hrishikesh Das / Tim Schulte / Ainhoa Moliner-Morro / Martin Corcoran / Adnane Achour / Gunilla B Karlsson Hedestam / B Martin ...Authors: Leo Hanke / Laura Vidakovics Perez / Daniel J Sheward / Hrishikesh Das / Tim Schulte / Ainhoa Moliner-Morro / Martin Corcoran / Adnane Achour / Gunilla B Karlsson Hedestam / B Martin Hällberg / Ben Murrell / Gerald M McInerney /
Abstract: SARS-CoV-2 enters host cells through an interaction between the spike glycoprotein and the angiotensin converting enzyme 2 (ACE2) receptor. Directly preventing this interaction presents an attractive ...SARS-CoV-2 enters host cells through an interaction between the spike glycoprotein and the angiotensin converting enzyme 2 (ACE2) receptor. Directly preventing this interaction presents an attractive possibility for suppressing SARS-CoV-2 replication. Here, we report the isolation and characterization of an alpaca-derived single domain antibody fragment, Ty1, that specifically targets the receptor binding domain (RBD) of the SARS-CoV-2 spike, directly preventing ACE2 engagement. Ty1 binds the RBD with high affinity, occluding ACE2. A cryo-electron microscopy structure of the bound complex at 2.9 Å resolution reveals that Ty1 binds to an epitope on the RBD accessible in both the 'up' and 'down' conformations, sterically hindering RBD-ACE2 binding. While fusion to an Fc domain renders Ty1 extremely potent, Ty1 neutralizes SARS-CoV-2 spike pseudovirus as a 12.8 kDa nanobody, which can be expressed in high quantities in bacteria, presenting opportunities for manufacturing at scale. Ty1 is therefore an excellent candidate as an intervention against COVID-19.
#1: Journal: BioRxiv / Year: 2020
Title: An alpaca nanobody neutralizes SARS-CoV-2 by blocking receptor interaction
Authors: Hanke L / Vidakovics L / Sheward DJ / Schulte T / Moliner Morro A / Corcoran M / Achour A / Karlsson Hedestam G / Hallberg BM / Murrell B / McInerney GM
History
DepositionJul 30, 2020-
Header (metadata) releaseSep 23, 2020-
Map releaseSep 23, 2020-
UpdateSep 23, 2020-
Current statusSep 23, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 3
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6zxn
  • Surface level: 3
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_11526.map.gz / Format: CCP4 / Size: 299.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationComposite map of prefusion SARS-CoV-2 spike glycoprotein with a single receptor binding domain up. The spike protein is bound with three neutralizing nanobodies.
Voxel sizeX=Y=Z: 1.02 Å
Density
Contour LevelBy AUTHOR: 2.74 / Movie #1: 3
Minimum - Maximum-43.933025 - 77.14982
Average (Standard dev.)0.00055904285 (±0.9428028)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions428428428
Spacing428428428
CellA=B=C: 436.56 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.021.021.02
M x/y/z428428428
origin x/y/z0.0000.0000.000
length x/y/z436.560436.560436.560
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ428428428
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS428428428
D min/max/mean-43.93377.1500.001

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Supplemental data

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Sample components

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Entire : Ternary complex between Nanobody Ty1 and Spike.

EntireName: Ternary complex between Nanobody Ty1 and Spike.
Components
  • Complex: Ternary complex between Nanobody Ty1 and Spike.
    • Complex: Nanobody strongly neutralising and blocks ACE2 binding.
      • Protein or peptide: Spike glycoproteinSpike protein
    • Complex: Nanobody Ty1
      • Protein or peptide: Nanobody Ty1
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: Ternary complex between Nanobody Ty1 and Spike.

SupramoleculeName: Ternary complex between Nanobody Ty1 and Spike. / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Molecular weightTheoretical: 14 KDa

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Supramolecule #2: Nanobody strongly neutralising and blocks ACE2 binding.

SupramoleculeName: Nanobody strongly neutralising and blocks ACE2 binding.
type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2
Recombinant expressionOrganism: Homo sapiens (human)

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Supramolecule #3: Nanobody Ty1

SupramoleculeName: Nanobody Ty1 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Vicugna pacos (alpaca)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Macromolecule #1: Spike glycoprotein

MacromoleculeName: Spike glycoprotein / type: protein_or_peptide / ID: 1
Details: Severe acute respiratory syndrome coronavirus 2 spike glykoprotein
Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2
Molecular weightTheoretical: 142.399375 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MFVFLVLLPL VSSQCVNLTT RTQLPPAYTN SFTRGVYYPD KVFRSSVLHS TQDLFLPFFS NVTWFHAIHV SGTNGTKRFD NPVLPFNDG VYFASTEKSN IIRGWIFGTT LDSKTQSLLI VNNATNVVIK VCEFQFCNDP FLGVYYHKNN KSWMESEFRV Y SSANNCTF ...String:
MFVFLVLLPL VSSQCVNLTT RTQLPPAYTN SFTRGVYYPD KVFRSSVLHS TQDLFLPFFS NVTWFHAIHV SGTNGTKRFD NPVLPFNDG VYFASTEKSN IIRGWIFGTT LDSKTQSLLI VNNATNVVIK VCEFQFCNDP FLGVYYHKNN KSWMESEFRV Y SSANNCTF EYVSQPFLMD LEGKQGNFKN LREFVFKNID GYFKIYSKHT PINLVRDLPQ GFSALEPLVD LPIGINITRF QT LLALHRS YLTPGDSSSG WTAGAAAYYV GYLQPRTFLL KYNENGTITD AVDCALDPLS ETKCTLKSFT VEKGIYQTSN FRV QPTESI VRFPNITNLC PFGEVFNATR FASVYAWNRK RISNCVADYS VLYNSASFST FKCYGVSPTK LNDLCFTNVY ADSF VIRGD EVRQIAPGQT GKIADYNYKL PDDFTGCVIA WNSNNLDSKV GGNYNYLYRL FRKSNLKPFE RDISTEIYQA GSTPC NGVE GFNCYFPLQS YGFQPTNGVG YQPYRVVVLS FELLHAPATV CGPKKSTNLV KNKCVNFNFN GLTGTGVLTE SNKKFL PFQ QFGRDIADTT DAVRDPQTLE ILDITPCSFG GVSVITPGTN TSNQVAVLYQ DVNCTEVPVA IHADQLTPTW RVYSTGS NV FQTRAGCLIG AEHVNNSYEC DIPIGAGICA SYQTQTNSPG SASSVASQSI IAYTMSLGAE NSVAYSNNSI AIPTNFTI S VTTEILPVSM TKTSVDCTMY ICGDSTECSN LLLQYGSFCT QLNRALTGIA VEQDKNTQEV FAQVKQIYKT PPIKDFGGF NFSQILPDPS KPSKRSFIED LLFNKVTLAD AGFIKQYGDC LGDIAARDLI CAQKFNGLTV LPPLLTDEMI AQYTSALLAG TITSGWTFG AGAALQIPFA MQMAYRFNGI GVTQNVLYEN QKLIANQFNS AIGKIQDSLS STASALGKLQ DVVNQNAQAL N TLVKQLSS NFGAISSVLN DILSRLDPPE AEVQIDRLIT GRLQSLQTYV TQQLIRAAEI RASANLAATK MSECVLGQSK RV DFCGKGY HLMSFPQSAP HGVVFLHVTY VPAQEKNFTT APAICHDGKA HFPREGVFVS NGTHWFVTQR NFYEPQIITT DNT FVSGNC DVVIGIVNNT VYDPLQPELD SFKEELDKYF KNHTSPDVDL GDISGINASV VNIQKEIDRL NEVAKNLNES LIDL QELGK YEQGSGYIPE APRDGQAYVR KDGEWVLLST FLGRSLEVLF QGPGHHHHHH HHSAWSHPQF EKGGGSGGGG SGGSA WSHP QFEK

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Macromolecule #2: Nanobody Ty1

MacromoleculeName: Nanobody Ty1 / type: protein_or_peptide / ID: 2
Details: Nanobody Ty1 that neutralises SARS-CoV-2 by blocking the receptor bindings site on the spike glykoprotein
Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Vicugna pacos (alpaca)
Molecular weightTheoretical: 14.306924 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MAQVQLVETG GGLVQPGGSL RLSCAASGFT FSSVYMNWVR QAPGKGPEWV SRISPNSGNI GYTDSVKGRF TISRDNAKNT LYLQMNNLK PEDTALYYCA IGLNLSSSSV RGQGTQVTVS SGGLPETGGH HHHHH

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 39 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4
GridMaterial: NICKEL/TITANIUM / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 20.0 µm / Calibrated defocus min: 0.3 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.3 µm / Nominal magnification: 165000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 10 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average exposure time: 1.5 sec. / Average electron dose: 51.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: Warp (ver. 1.0.7)
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 2.93 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15) / Number images used: 131816
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:
RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-6zxn:
Cryo-EM structure of the SARS-CoV-2 spike protein bound to neutralizing nanobodies (Ty1)

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